A risk-reward examination of sample multiplexing reagents for single cell RNA-Seq.

Single-cell RNA sequencing (scRNA-Seq) has emerged as a powerful tool for understanding cellular heterogeneity and function. However the choice of sample multiplexing reagents can impact data quality and experimental outcomes. In this study, we compared various multiplexing reagents, including MULTI-Seq, Hashtag antibody, and CellPlex, across diverse sample types such as human peripheral blood mononuclear cells (PBMCs), mouse embryonic brain and patient-derived xenografts (PDXs). We found that all multiplexing reagents worked well in cell types robust to ex vivo manipulation but suffered from signal-to-noise issues in more delicate sample types. We compared multiple demultiplexing algorithms which differed in performance depending on data quality. We find that minor improvements to laboratory workflows such as titration and rapid processing are critical to optimal performance. We also compared the performance of fixed scRNA-Seq kits and highlight the advantages of the Parse Biosciences kit for fragile samples. Highly multiplexed scRNA-Seq experiments require more sequencing resources, therefore we evaluated CRISPR-based destruction of non-informative genes to enhance sequencing value. Our comprehensive analysis provides insights into the selection of appropriate sample multiplexing reagents and protocols for scRNA-Seq experiments, facilitating more accurate and cost-effective studies.

A perioperative study of Safusidenib in patients with IDH1-mutated glioma.

This is a single arm, open label perioperative trial to assess the feasibility, pharmacokinetics and pharmacodynamics of treatment with safusidenib following biopsy, and prior to surgical resection in patients with IDH1 mutated glioma who have not received radiation therapy or chemotherapy. Fifteen participants will receive treatment in two parts. First, biopsy followed by one cycle (28 days) of safusidenib, an orally available, small molecular inhibitor of mutated IDH1, then maximal safe resection of the tumor (Part A). Second, after recovery from surgery, safusidenib until disease progression or unacceptable toxicity (Part B). This research will enable objective measurement of biological activity of safusidenib in patients with IDH1 mutated glioma. Anti-tumor activity will be assessed by progression free survival and time to next intervention.Clinical Trial Registration: NCT05577416 (ClinicalTrials.gov).

Adult low-grade gliomas (aLGG) are primary brain cancers, defined by mutations in IDH1 or IDH2. When the IDH gene becomes abnormal (mutated), production of a metabolite that causes cancer cells to grow is increased. These tumors grow slowly but invade the normal functioning brain, making them nearly impossible to cure. The current standard of care treatment includes surgery, followed by radiation therapy and chemotherapy, the timing of which depends on the risk of cancer regrowth. Some patients may be suitable for monitoring with MRI scans alone, however recurrences will inevitably occur. Recently developed targeted mutant IDH inhibitors for aLGG patients may be beneficial both at diagnosis and recurrence. Notably, early treatment prior to radiation therapy and chemotherapy delays growth of aLGG and the need for subsequent radiation therapy and chemotherapy. Nevertheless, most patients will eventually suffer further tumor growth and the optimal timing and sequencing of these therapies remains an area of active research. This research investigates the mutant IDH1 inhibitor safusidenib. The researchers are conducting an innovative clinical trial where patients with aLGG, who have not received radiation therapy or chemotherapy, are treated with safusidenib following a biopsy and prior to surgical removal of their tumor. In this study they investigate whether this trial design is safe and feasible, and how safusidenib works; with the goal to better understand the optimal use of IDH inhibitors for patients with aLGG.

A Multiplexed Approach to Assess Small Cell Lung Cancer Subtype Heterogeneity in Primary and Patient-Derived Tumor Samples.

Unlocking the heterogeneity of cancers is crucial for developing therapeutic approaches that effectively eradicate disease. As our understanding of markers specific to cancer subclones or subtypes expands, there is a growing demand for advanced technologies that enable the simultaneous investigation of multiple targets within an individual tumor sample. Indeed, multiplex approaches offer distinct benefits, particularly when tumor specimens are small and scarce. Here we describe the utility of two fluorescence-based multiplex approaches; fluorescent Western blots, and multiplex immunohistochemistry (Opal™) staining to interrogate heterogeneity, using small cell lung cancer as an example. Critically, the coupling of Opal™ staining with advanced image quantitation, permits the dissection of cancer cell phenotypes at a single cell level. These approaches can be applied to patient biopsies and/or patient-derived xenograft (PDX) models and serve as powerful methodologies for assessing tumor cell heterogeneity in response to therapy or between metastatic lesions across diverse tissue sites.

The Landscape of Pediatric High-Grade Gliomas: The Virtues and Pitfalls of Pre-Clinical Models.

Pediatric high-grade gliomas (pHGG) are malignant and usually fatal central nervous system (CNS) WHO Grade 4 tumors. The majority of pHGG consist of diffuse midline gliomas (DMG), H3.3 or H3.1 K27 altered, or diffuse hemispheric gliomas (DHG) (H3.3 G34-mutant). Due to diffuse tumor infiltration of eloquent brain areas, especially for DMG, surgery has often been limited and chemotherapy has not been effective, leaving fractionated radiation to the involved field as the current standard of care. pHGG has only been classified as molecularly distinct from adult HGG since 2012 through Next-Generation sequencing approaches, which have shown pHGG to be epigenetically regulated and specific tumor sub-types to be representative of dysregulated differentiating cells. To translate discovery research into novel therapies, improved pre-clinical models that more adequately represent the tumor biology of pHGG are required. This review will summarize the molecular characteristics of different pHGG sub-types, with a specific focus on histone K27M mutations and the dysregulated gene expression profiles arising from these mutations. Current and emerging pre-clinical models for pHGG will be discussed, including commonly used patient-derived cell lines and in vivo modeling techniques, encompassing patient-derived xenograft murine models and genetically engineered mouse models (GEMMs). Lastly, emerging techniques to model CNS tumors within a human brain environment using brain organoids through co-culture will be explored. As models that more reliably represent pHGG continue to be developed, targetable biological and genetic vulnerabilities in the disease will be more rapidly identified, leading to better treatments and improved clinical outcomes.